阅读说明：本技术 新型人工核酸分子 (Novel artificial nucleic acid molecules ) 是由 托马斯·施拉克 安德烈亚斯·特斯 莫里茨·特兰 弗雷德里克·切韦西耶-通纳森 马里翁·波尼施 于 2018-10-17 设计创作，主要内容包括：本发明提供了包含5′非翻译区(UTR)元件和3′UTR元件的新组合的人工核酸分子。本发明的核酸分子优选地以提高可操作地连接至所述UTR元件的编码区的表达效率为特征。人工核酸可用于治疗或预防各种疾病。本发明还提供了包含所述人工核酸分子的(药物)组合物、疫苗和试剂盒。此外,提供了制备根据本发明的人工核酸分子的体外方法。(The present invention provides artificial nucleic acid molecules comprising a novel combination of 5 'untranslated region (UTR) elements and 3' UTR elements. The nucleic acid molecules of the invention are preferably characterized by an increased expression efficiency of a coding region operably linked to said UTR element. The artificial nucleic acid can be used for treating or preventing various diseases. The invention also provides (pharmaceutical) compositions, vaccines and kits comprising said artificial nucleic acid molecule. Furthermore, an in vitro method for the preparation of the artificial nucleic acid molecule according to the invention is provided.)

1. An artificial nucleic acid molecule comprising

a. At least one 5 ' untranslated region (5 ' UTR) element derived from a 5 ' UTR of a gene selected from HSD17B4, ASAH1, ATP5a1, MP68, ndifa 4, NOSIP, RPL31, SLC7A3, TUBB4B, and UBQLN 2;

b. at least one 3 ' untranslated region (3 ' UTR) element derived from a3 ' UTR of a gene selected from PSMB3, CASP1, COX6B1, GNAS, ndifa 1, and RPS 9; and optionally

c. At least one coding region operably linked to the 5 'UTR and the 3' UTR.

2. The artificial nucleic acid molecule of claim 1, wherein the 5 'UTR and/or the 3' UTR are heterologous to the coding region.

3. The artificial nucleic acid molecule of any one of claims 1 or 2, wherein each UTR comprises a naturally occurring DNA sequence and homologs, variants, fragments, and corresponding RNA sequences thereof.

4. The artificial nucleic acid molecule according to any one of claims 1 to 3, comprising

a-1. at least one 5 'UTR element derived from the 5' UTR of HSD17B4 gene or its corresponding RNA sequence, homologue, fragment or variant, and at least one 3 'UTR element derived from the 3' UTR of PSMB3 gene or its corresponding RNA sequence, homologue, fragment or variant; or

a-2. at least one 5 'UTR element derived from the 5' UTR of the ndifa 4 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the PSMB3 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

a-3. at least one 5 'UTR element derived from the 5' UTR of the SLC7a3 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the PSMB3 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

a-4. at least one 5 'UTR element derived from the 5' UTR of the NOSIP gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the PSMB3 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

a-5. at least one 5 'UTR element derived from the 5' UTR of the MP68 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the PSMB3 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

b-1. at least one 5 'UTR element derived from the 5' UTR of the UBQLN2 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the RPS9 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

At least one 5 'UTR element derived from the 5' UTR of the ASAH1 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the RPS9 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

At least one 5 'UTR element derived from the 5' UTR of HSD17B4 gene or its corresponding RNA sequence, homologue, fragment or variant, and at least one 3 'UTR element derived from the 3' UTR of RPS9 gene or its corresponding RNA sequence, homologue, fragment or variant; or

At least one 5 'UTR element derived from the 5' UTR of HSD17B4 gene or its corresponding RNA sequence, homologue, fragment or variant, and at least one 3 'UTR element derived from the 3' UTR of CASP1 gene or its corresponding RNA sequence, homologue, fragment or variant; or

At least one 5 'UTR element derived from the 5' UTR of the NOSIP gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the COX6B1 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

c-1. at least one 5 'UTR element derived from the 5' UTR of the ndifa 4 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the RPS9 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

c-2. at least one 5 'UTR element derived from the 5' UTR of the NOSIP gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the ndifa 1 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

c-3. at least one 5 'UTR element derived from the 5' UTR of the ndifa 4 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the COX6B1 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

c-4. at least one 5 'UTR element derived from the 5' UTR of the ndifa 4 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the ndifa 1 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

c-5. at least one 5 'UTR element derived from the 5' UTR of the ATP5a1 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the PSMB3 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

d-1. at least one 5 'UTR element derived from the 5' UTR of the RPL31 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the PSMB3 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

At least one 5 'UTR element derived from the 5' UTR of the ATP5a1 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the CASP1 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

At least one 5 'UTR element derived from the 5' UTR of the SLC7a3 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the GNAS1 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

d-4. at least one 5 'UTR element derived from the 5' UTR of HSD17B4 gene or its corresponding RNA sequence, homologue, fragment or variant, and at least one 3 'UTR element derived from the 3' UTR of NDUFA1 gene or its corresponding RNA sequence, homologue, fragment or variant; or

At least one 5 'UTR element derived from the 5' UTR of the SLC7A3 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the NDUFA1 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

e-1. at least one 5 'UTR element derived from the 5' UTR of the TUBB4B gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the RPS9 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

e-2. at least one 5 'UTR element derived from the 5' UTR of the RPL31 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the RPS9 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

At least one 5 'UTR element derived from the 5' UTR of the MP68 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the RPS9 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

At least one 5 'UTR element derived from the 5' UTR of the NOSIP gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the RPS9 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

e-5. at least one 5 'UTR element derived from the 5' UTR of the ATP5a1 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the RPS9 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

At least one 5 'UTR element derived from the 5' UTR of the ATP5a1 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the COX6B1 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

f-1. at least one 5 'UTR element derived from the 5' UTR of the ATP5a1 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the GNAS gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

f-2. at least one 5 'UTR element derived from the 5' UTR of the ATP5a1 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the ndifa 1 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

At least one 5 'UTR element derived from the 5' UTR of the HSD17B4 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the COX6B1 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

f-4. at least one 5 'UTR element derived from the 5' UTR of HSD17B4 gene or its corresponding RNA sequence, homologue, fragment or variant, and at least one 3 'UTR element derived from the 3' UTR of GNAS1 gene or its corresponding RNA sequence, homologue, fragment or variant; or

f-5. at least one 5 'UTR element derived from the 5' UTR of the MP68 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the COX6B1 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

g-1. at least one 5 'UTR element derived from the 5' UTR of MP68 gene or its corresponding RNA sequence, homologue, fragment or variant, and at least one 3 'UTR element derived from the 3' UTR of NDUFA1 gene or its corresponding RNA sequence, homologue, fragment or variant; or

g-2. at least one 5 'UTR element derived from the 5' UTR of the ndifa 4 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the CASP1 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

g-3. at least one 5 'UTR element derived from the 5' UTR of the ndifa 4 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the GNAS gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

g-4. at least one 5 'UTR element derived from the 5' UTR of the NOSIP gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the CASP1 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

g-5. at least one 5 'UTR element derived from the 5' UTR of the RPL31 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the CASP1 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

h-1. at least one 5 'UTR element derived from the 5' UTR of the RPL31 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the COX6B1 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

h-2. at least one 5 'UTR element derived from the 5' UTR of the RPL31 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the GNAS gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

h-3. at least one 5 'UTR element derived from the 5' UTR of the RPL31 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the ndifa 1 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

h-4. at least one 5 'UTR element derived from the 5' UTR of the SLC7a3 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the CASP1 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

h-5. at least one 5 'UTR element derived from the 5' UTR of the SLC7A3 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the COX6B1 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

i-1. at least one 5 'UTR element derived from the 5' UTR of the SLC7a3 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the RPS9 gene or a corresponding RNA sequence, homologue, fragment or variant thereof; or

i-2. at least one 5 'UTR element derived from the 5' UTR of the ndefa4.1 gene or a corresponding RNA sequence, homologue, fragment or variant thereof, and at least one 3 'UTR element derived from the 3' UTR of the CASP1 gene or a corresponding RNA sequence, homologue, fragment or variant thereof.

5. The artificial nucleic acid molecule according to claim 4, comprising a UTR element according to a-1, a-2, a-3, a-4 or a-5, preferably a UTR element according to a-1.

6. The artificial nucleic acid molecule according to claim 4, comprising a nucleic acid molecule according to a-2(NDUFA4/PSMB 3); a-5(MP68/PSMB 3); c-1(NDUFA4/RPS 9); a-1(HSD17B4/PSMB 3); e-3(MP68/RPS 9); e-4(NOSIP/RPS 9); a-4(NOSIP/PSMB 3); e-2(RPL31/RPS 9); e-5(ATP5A1/RPS 9); d-4(HSD17B4/NUDFA 1); b-5(NOSIP/COX6B 1); a-3(SLC7A3/PSMB 3); b-1(UBQLN2/RPS 9); b-2(ASAH1/RPS 9); b-4(HSD17B4/CASP 1); e-6(ATP5A1/COX6B 1); b-3(HSD17B4/RPS 9); g-5(RPL31/CASP 1); h-1(RPL31/COX6B 1); and/or UTR elements of c-5(ATP5A1/PSMB 3).

7. The artificial nucleic acid molecule according to claim 4, comprising a nucleic acid molecule according to a-1(HSD17B4/PSMB 3); a-3(SLC7A3/PSMB 3); e-2(RPL31/RPS 9); a-5(MP68/PSMB 3); d-1(RPL31/PSMB 3); a-2(NDUFA4/PSMB 3); h-1(RPL31/COX6B 1); b-1(UBQLN2/RPS 9); a-4(NOSIP/PSMB 3); c-5(ATP5A1/PSMB 3); b-5(NOSIP/COX6B 1); d-4(HSD17B4/NDUFA 1); i-1(SLC7A3/RPS 9); i-2(NduFa4.1/CASP 1); f-3(HSD17B4/COX6B 1); b-4(HSD17B4/CASP 1); g-5(RPL31/CASP 1); c-2(NOSIP/NDUFA 1); e-4(NOSIP/RPS 9); c-4(NDUFA4/NDUFA 1); and/or d-5(SLC7A3/NDUFA 1).

8. The artificial nucleic acid molecule according to claim 4, comprising a nucleic acid molecule according to a-4(NOSIP/PSMB 3); a-1(HSD17B4/PSMB 3); a-5(MP68/PSMB 3); d-3(SLC7A 3/GNAS); a-2(NDUFA4/PSMB 3); a-3(SLC7A3/PSMB 3); d-5(SLC7A3/NDUFA 1); i-1(SLC7A3/RPS 9); d-1(RPL31/PSMB 3); d-4(HSD17B4/NDUFA 1); b-3(HSD17B4/RPS 9); f-3(HSD17B4/COX6B 1); f-4(HSD17B 4/GNAS); h-5(SLC7A3/COX6B 1); g-4(NOSIP/CASP 1); c-3(NDUFA4/COX6B 1); b-1(UBQLN2/RPS 9); c-5(ATP5A1/PSMB 3); h-4(SLC7A3/CASP 1); h-2(RPL 31/GNAS); e-1(TUBB4B/RPS 9); f-2(ATP5A1/NDUFA 1); c-2(NOSIP/NDUFA 1); b-5(NOSIP/COX6B 1); and/or UTR elements of e-4(NOSIP/RPS 9.1).

9. The artificial nucleic acid molecule according to any one of claims 1 to 8, wherein

-the 5' UTR element derived from HSD17B4 gene comprises or consists of a sequence according to SEQ ID NO: 1, or a DNA sequence according to SEQ ID NO: 1 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority; or according to SEQ id no: 2, or an RNA sequence according to SEQ ID NO: 2 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority;

-the 5' UTR element derived from the ASAH1 gene comprises or consists of a sequence according to SEQ ID NO: 3, or a DNA sequence according to SEQ ID NO: 3 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority; or according to SEQ ID NO: 4, or an RNA sequence according to SEQ ID NO: 4 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority;

-the 5' UTR element derived from the ATP5a1 gene comprises or consists of a sequence according to SEQ ID NO: 5, or a DNA sequence according to SEQ ID NO: 5 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority; or according to SEQ ID NO: 6, or an RNA sequence according to SEQ ID NO: 6 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority;

-the 5' UTR element derived from the MP68 gene comprises or consists of a sequence according to SEQ ID NO: 7, or a DNA sequence according to SEQ ID NO: 7 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority; or according to SEQ ID NO: 8, or an RNA sequence according to SEQ ID N0: 8 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority;

-the 5' UTR element derived from the ndifa 4 gene comprises or consists of a sequence according to SEQ ID NO: 9, or a DNA sequence according to SEQ ID NO: 9 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority; or according to SEQ ID NO: 10, or an RNA sequence according to SEQ ID NO: 10 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority;

-the 5' UTR element derived from the NOSIP gene comprises or consists of a sequence according to SEQ ID NO: 11, or a DNA sequence according to SEQ ID NO: 11 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority; or according to SEQ id no: 12, or an RNA sequence according to SEQ ID NO: 12 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority;

-the 5' UTR element derived from the RPL31 gene comprises or consists of a sequence according to SEQ ID NO: 13, or a DNA sequence according to SEQ ID NO: 13 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority; according to SEQ ID NO: 14, or an RNA sequence according to SEQ ID NO: 14 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority;

-the 5' UTR element derived from the SLC7a3 gene comprises or consists of a sequence according to SEQ ID NO: 15, or a DNA sequence according to SEQ ID NO: 15 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority; or according to SEQ id no: 16, or an RNA sequence according to SEQ ID NO: 16 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority;

-the 5' UTR element derived from the TUBB4B gene comprises or consists of a sequence according to SEQ ID NO: 17, or a DNA sequence according to SEQ ID NO: 17 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority; or according to SEQ id no: 18, or an RNA sequence according to SEQ ID NO: 18 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority;

-the 5' UTR element derived from the UBQLN2 gene comprises or consists of a sequence according to SEQ ID NO: 19, or a DNA sequence according to SEQ ID NO: 19 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority; or according to SEQ id no: 20, or an RNA sequence according to SEQ ID NO: 20 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority;

-the 3' UTR element derived from the PSMB3 gene comprises or consists of a sequence according to SEQ ID NO: 23, or a DNA sequence according to SEQ ID NO: 23 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority; or according to SEQ id no: 24, or an RNA sequence according to SEQ ID NO: 24 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority;

-the 3' UTR element derived from the CASP1 gene comprises or consists of a sequence according to SEQ ID NO: 25, or a DNA sequence according to SEQ ID NO: 25 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority; or according to SEQ id no: 26, or an RNA sequence according to SEQ ID NO: 26 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority;

-the 3' UTR element derived from the COX6B1 gene comprises or consists of the amino acid sequence according to SEQ ID NO: 27, or a DNA sequence according to SEQ ID NO: 27 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority; or according to SEQ id no: 28, or an RNA sequence according to SEQ ID NO: 28, or a fragment or variant thereof, having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority;

-the 3' UTR element derived from the GNAS gene comprises or consists of a sequence according to SEQ ID NO: 29, or a DNA sequence according to SEQ ID NO: 29 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority; or according to SEQ id no: 30, or an RNA sequence according to SEQ ID NO: 30 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority;

-the 3' UTR element derived from the ndifa 1 gene comprises or consists of a sequence according to SEQ ID NO: 31, or a DNA sequence according to SEQ ID NO: 31 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority; or according to SEQ id no: 32, or an RNA sequence according to SEQ ID NO: 32, or a fragment or variant thereof, having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority; and/or

-the 3' UTR element derived from the RPS9 gene comprises or consists of a sequence according to SEQ ID NO: 33, or a DNA sequence according to SEQ ID NO: 33 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority; or according to SEQ id no: 34, or an RNA sequence according to SEQ ID NO: 34 or a fragment or variant thereof having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending order of priority.

10. The artificial nucleic acid molecule according to any one of claims 1 to 9, wherein the coding region is located between the 5 'UTR and the 3' UTR, preferably downstream of the 5 'UTR and upstream of the 3' UTR.

11. The artificial nucleic acid molecule according to any one of claims 1 to 10, wherein the at least one coding region encodes at least one (poly) peptide or protein of interest, optionally selected from an antigenic (poly) peptide or protein, an allergenic (poly) peptide or protein, a therapeutic (poly) peptide or protein, an antibody or a fragment, variant or derivative of said (poly) peptide or protein of interest.

12. The artificial nucleic acid molecule according to claim 11, wherein at least one antigenic (poly) peptide or protein is selected from a tumor antigen, a pathogenic antigen, an autoantigen, an alloantigen or an allergenic antigen.

13. The artificial nucleic acid molecule according to claim 12, wherein at least one pathogenic antigen is selected from a bacterial antigen, a viral antigen, a fungal antigen or a protozoan antigen.

14. The artificial nucleic acid molecule according to claim 11, wherein the therapeutic (poly) peptide or protein is selected from

-a therapeutic (poly) peptide or protein that replaces a deleted, defective or mutated protein;

therapeutic (poly) peptides or proteins useful for the treatment of genetic or acquired diseases, infectious diseases or tumors (e.g. cancer or tumor diseases);

-a secondary or immunostimulatory therapeutic (poly) peptide or protein;

-a therapeutic antibody;

-a peptide hormone;

-a gene editing agent;

-an immune checkpoint inhibitor;

-a T cell receptor;

-an enzyme; and/or

-a variant, fragment or derivative of any of said therapeutic (poly) peptides or proteins.

15. The artificial nucleic acid molecule according to any one of claims 10 to 14, wherein the at least one coding region further encodes

(a) At least one effector domain;

(b) at least one peptide tag or protein tag;

(c) at least one localization signal or localization sequence;

(d) at least one Nuclear Localization Signal (NLS);

(e) at least one signal peptide; and/or

(f) At least one peptide linker;

(g) a Secretion Signal Peptide (SSP),

(h) a multimerization element comprising a dimerization element, a trimerization element, a tetramerization element, or an oligomerization element;

(i) a Virus Like Particle (VLP) forming element;

(j) a transmembrane element;

(k) a dendritic cell targeting element;

(l) An immune accessory element;

(m) an element that facilitates antigen presentation;

(n) a 2A peptide;

(o) an element that extends the half-life of the protein; and/or

(p) elements for post-translational modification (e.g., glycosylation),

wherein the artificial nucleic acid molecule further optionally comprises at least one Internal Ribosome Entry Site (IRES) and/or at least one miRNA binding site.

16. The artificial nucleic acid molecule according to any one of claims 1 to 15, wherein the at least one coding region encodes a polypeptide comprising or consisting of a sequence according to SEQ ID NO: 41 to 45, or to an amino acid sequence according to any one of SEQ ID NOs: 42 to 45, or a variant or fragment of any of these sequences, in ascending order of priority.

17. The artificial nucleic acid molecule according to any one of claims 1 to 15, wherein the at least one coding region of the artificial nucleic acid molecule comprises or consists of a sequence according to SEQ ID NO: 46 to 49, or a nucleic acid sequence having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending priority order with any of said nucleic acid sequences.

18. The artificial nucleic acid molecule according to any one of claims 1 to 16, wherein the artificial nucleic acid molecule comprises or consists of an amino acid sequence according to SEQ ID NO: 50 to 368, or a nucleic acid sequence having at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity in ascending priority order with any of said nucleic acid sequences.

19. The artificial nucleic acid molecule according to any one of claims 1 to 17, wherein the artificial nucleic acid molecule is RNA.

20. The RNA of claim 19, wherein the RNA is a monocistronic RNA, a bicistronic RNA, or a polycistronic RNA.

21. The RNA according to claim 19 or 20, wherein the RNA is mRNA, viral RNA, self-replicating RNA, or replicon RNA.

22. The artificial nucleic acid, preferably RNA, according to any one of claims 1 to 21, wherein the artificial nucleic acid is a modified nucleic acid, preferably a stabilized nucleic acid, or wherein the artificial nucleic acid comprises at least one modified or non-naturally occurring nucleotide, backbone modification, sugar modification or base modification.

23. The artificial nucleic acid, preferably RNA, according to any one of claims 1 to 22, wherein

-the G/C content of at least one coding region of the artificial nucleic acid is increased compared to the G/C content of the corresponding coding sequence of the corresponding wild-type artificial nucleic acid, and/or wherein

-the C content of at least one coding region of an artificial nucleic acid is increased compared to the C content of the corresponding coding sequence of the corresponding wild-type artificial nucleic acid, and/or wherein

-codons in at least one coding region of the artificial nucleic acid are adapted to human codon usage, wherein a Codon Adaptation Index (CAI) is preferably increased or maximized in the at least one coding sequence of the artificial nucleic acid,

-wherein the amino acid sequence encoded by the artificial nucleic acid is preferably not altered compared to the amino acid sequence encoded by the corresponding wild-type artificial nucleic acid.

24. The artificial nucleic acid, preferably RNA, according to any one of claims 1 to 23, comprising a 5' -cap structure, preferably m7GpppN or cap 1.

25. The artificial nucleic acid, preferably RNA, according to any one of claims 1 to 24, comprising at least one histone stem-loop.

26. The artificial nucleic acid, preferably RNA, according to claim 25, wherein the at least one histone stem-loop comprises a nucleic acid sequence according to the following formula (I) or (II):

formula (I) (stem-loop sequence without stem-adjoining elements):

formula (II) (stem-loop sequence with stem-adjoining elements):

wherein:

stem 1 adjoining element or stem 2 adjoining element N_1-6A nucleotide or nucleotide analogue thereof which is a contiguous sequence of 1 to 6, preferably 2 to 6, more preferably 2 to 5, even more preferably 3 to 5, most preferably 4 to 5 or 5N, wherein each N is independently from each other selected from A, U, T, G and C;

stem 1[ N ]_0-2GN_3-5]Is reverse complementary or partially reverse complementary to element stem 2 and is a contiguous sequence of 5 to 7 nucleotides;

wherein N is_0-2A nucleotide or nucleotide analogue thereof which is a contiguous sequence of 0 to 2, preferably 0 to 1, more preferably 1N, wherein each N is independently from each other selected from A, U, T, G and C;

wherein N is_3-5Is a contiguous sequence of 3 to 5, preferably 4 to 5, more preferably 4, N, wherein each N is independently from each other a nucleotide or nucleotide analogue thereof selected from the group consisting of A, U, T, G and C, and

wherein G is guanosine or an analog thereof, and may optionally be replaced by cytidine or an analog thereof, provided that its complementary nucleotide in stem 2, cytidine, is replaced by guanosine;

loop sequence [ N ]_0-4(U/T)N_0-4]Located between element stem 1 and stem 2 and is a continuous sequence of 3 to 5 nucleotides, more preferably 4 nucleotides;

wherein each N_0-4A contiguous sequence of 0 to 4, preferably 1 to 3, more preferably 1 to 2N, independently of each other, wherein each N is independently of each other selected from A, U, T, G and C nucleotides or nucleotide analogues thereof; and

wherein U/T represents uridine, or optionally thymidine;

stem 2[ N ]_3-5CN_0-2]Is reverse complementary or partially reverse complementary to element stem 1 and is a contiguous sequence of 5 to 7 nucleotides;

wherein N is_3-5A nucleotide or nucleotide analogue thereof which is a contiguous sequence of 3 to 5, preferably 4 to 5, more preferably 4N, wherein each N is independently from each other selected from A, U, T, G and C;

wherein N is_0-2A nucleotide or nucleotide analogue thereof which is a contiguous sequence of 0 to 2, preferably 0 to 1, more preferably 1N, wherein each N is independently from each other selected from A, U, T, G and C; and

wherein C is cytidine or an analog thereof, and may optionally be replaced by guanosine or an analog thereof, provided that its complementary nucleotide in stem 1, guanosine, is replaced by cytidine;

wherein

Stem 1 and stem 2 are capable of base pairing with each other,

form a reverse complementary sequence in which base pairing can occur between stem 1 and stem 2, or

A partially reverse complementary sequence is formed in which incomplete base pairing can occur between stem 1 and stem 2.

27. The artificial nucleic acid, preferably RNA, according to claim 25 or 26, wherein the at least one histone stem-loop comprises a nucleic acid sequence according to the following formula (Ia) or (IIa):

formula (Ia) (stem-loop sequence without stem-adjoining elements):

formula (IIa) (stem-loop sequence with stem-adjacent elements):

28. the artificial nucleic acid, preferably RNA, according to any one of claims 1 to 27, optionally comprising a polyadenylation sequence, preferably comprising 10 to 200, 10 to 100, 40 to 80, or 50 to 70 adenine nucleotides.

29. The artificial nucleic acid, preferably RNA, according to any one of claims 1 to 28, optionally comprising a polycytidylic acid sequence, preferably comprising 10 to 200, 10 to 100, 20 to 70, 20 to 60, or 10 to 40 cytosine nucleotides.

30. The artificial nucleic acid, preferably RNA, according to any one of claims 1 to 29, comprising, preferably in the 5 'to 3' direction, the following elements:

a) a 5' -cap structure, preferably m7GpppN or cap 1;

b) a 5 '-UTR element comprising or consisting of a nucleic acid sequence derived from a 5' -UTR as defined in any of claims 1 to 9, preferably comprising a sequence corresponding to the sequence according to SEQ ID NO: 1 to 20, or a homologue, fragment or variant thereof;

c) at least one coding sequence as defined in any one of claims 10 to 18,

d) a3 '-UTR element comprising or consisting of a nucleic acid sequence derived from a 3' -UTR as defined in any of claims 1 to 9, preferably comprising a sequence corresponding to the sequence according to SEQ ID NO: 23 to 34, or a homologue, fragment or variant thereof;

e) optionally a poly-A tail, which preferably consists of 10 to 1000, 10 to 500, 10 to 300, 10 to 200, 10 to 100, 40 to 80 or 50 to 70 adenine nucleotides,

f) optionally a poly-cytosine tail, which preferably consists of 10 to 200, 10 to 100, 20 to 70, 20 to 60 or 10 to 40 cytosine nucleotides, and

g) optionally a histone stem-loop.

31. A composition comprising at least one or more artificial nucleic acid molecules, preferably RNA, according to any one of claims 1 to 30, and a pharmaceutically acceptable carrier and/or excipient.

32. The composition of claim 31, wherein at least two of the plurality of artificial nucleic acid molecules each (a) comprise the same or different combination of UTR elements according to any one of claims 1 to 9, and/or (b) encode a different peptide or protein, optionally selected from the group consisting of peptides or proteins according to any one of claims 11 to 17.

33. A composition according to claim 31 or 32 for use as a medicament, optionally as a vaccine.

34. The (pharmaceutical) composition according to claim 33, preferably comprising at least one artificial nucleic acid molecule comprising a UTR combination according to claim 6, wherein the (pharmaceutical) composition and/or the artificial nucleic acid molecule is suitable for liver-targeted delivery.

35. The (pharmaceutical) composition according to claim 33, preferably comprising at least one artificial nucleic acid molecule comprising a UTR combination according to claim 7, wherein the (pharmaceutical) composition and/or the artificial nucleic acid molecule is suitable for subcutaneous, intradermal, topical or transdermal administration.

36. The (pharmaceutical) composition according to claim 33, preferably comprising at least one artificial nucleic acid molecule comprising a UTR combination according to claim 8, wherein the (pharmaceutical) composition and/or the artificial nucleic acid molecule is suitable for intramuscular administration.

37. The (pharmaceutical) composition or vaccine according to any one of claims 31 to 36, wherein the artificial nucleic acid molecule, preferably RNA, is complexed with one or more than one cationic or polycationic compound, preferably a cationic or polycationic polymer, a cationic peptide or protein or a polycationic peptide or protein such as protamine, a cationic or polycationic polysaccharide and/or a cationic lipid or a polymeric carrier or a polycationic lipid or a polymeric carrier.

38. The (pharmaceutical) composition or vaccine according to claim 37, wherein the N/P ratio of the artificial nucleic acid molecule, preferably RNA, to one or more cationic peptides or proteins or polycationic peptides or proteins is about 0.1 to 10, including about 0.3 to 4, about 0.5 to 2, about 0.7 to 2 and about 0.7 to 1.5.

39. The (pharmaceutical) composition or vaccine according to any one of claims 31 to 38, wherein the artificial nucleic acid molecule, preferably RNA, is complexed with one or more than one lipid, thereby forming lipid nanoparticles, lipid complexes and/or preferably liposomes.

40. The (pharmaceutical) composition or vaccine according to any one of claims 31 to 39, further comprising at least one further active agent and/or at least one adjuvant.

41. The (pharmaceutical) composition or vaccine of any one of claims 31 to 40, further comprising non-coding RNAs selected from the group consisting of small interfering RNAs (siRNAs), antisense RNAs (asRNAs), circular RNAs (circRNAs), ribozymes, aptamers, riboswitches, immunostimulatory RNAs (isRNAs), transfer RNAs (tRNAs), ribosomal RNAs (rRNAs), small intranuclear RNAs (snRNAs), small nucleolar RNAs (snornas), micro RNAs (miRNAs), and Piwi-interacting RNAs (piRNAs).

42. The (pharmaceutical) composition or vaccine of claim 41, wherein the immunostimulatory RNA (isRNA) comprises a (G) according to formula (III)_lX_mG_n) Formula (IV) (C)_lX_mC_n) Formula (V) (N)_uG_lX_mG_nN_v)_aAnd/or of the formula (VI) (N)_uC_lX_mC_nN_v)_aAt least one RNA sequence of (a).

43. The (pharmaceutical) composition or vaccine of any one of claims 41 or 42, comprising a polymeric carrier cargo complex formed from a polymeric carrier, preferably comprising disulfide-linked cationic peptides, preferably Cys-Arg12 and/or Cys-Arg12-Cys, and isRNA.

44. A kit, preferably a kit of parts, comprising an artificial nucleic acid molecule, preferably RNA, according to any one of claims 1 to 30, or a (pharmaceutical) composition or vaccine according to any one of claims 31 to 43, and optionally a liquid carrier and/or optionally technical instructions for information on the administration and dosage of the artificial nucleic acid molecule or (pharmaceutical) composition or vaccine.

45. The kit of claim 44, wherein the kit comprises lactated ringer's solution as a part.

46. The artificial nucleic acid molecule, preferably RNA, according to any one of claims 1 to 30, the (pharmaceutical) composition or vaccine according to any one of claims 31 to 43, or the kit according to claim 44 or 45, for use as a medicament.

47. The artificial nucleic acid molecule, preferably RNA, according to any one of claims 1 to 30, the (pharmaceutical) composition or vaccine according to any one of claims 31 to 43, or the kit according to claim 44 or 45 for use in the treatment of genetic diseases, cancer, infectious diseases, inflammatory diseases, (auto) immune diseases, allergies, and/or for use in gene therapy and/or immunomodulation.

48. The artificial nucleic acid molecule, preferably RNA, (pharmaceutical) composition or vaccine or kit for use according to claim 47, wherein said use comprises (a) administering said artificial nucleic acid molecule, preferably RNA, said (pharmaceutical) composition or vaccine, or said kit, to a patient in need thereof.

49. The artificial nucleic acid molecule, preferably RNA, according to any one of claims 6 to 30, the (pharmaceutical) composition or vaccine according to any one of claims 31 to 43, or the kit according to claim 44 or 45, for use in a method for increasing the efficiency of expression of the artificial nucleic acid molecule in liver tissue, hepatocytes or a hepatocyte line, the (pharmaceutical) composition or kit comprising at least one artificial nucleic acid molecule according to any one of claims 6 to 30.

50. The artificial nucleic acid molecule, preferably RNA, according to any one of claims 7 to 30, the (pharmaceutical) composition or vaccine according to any one of claims 31 to 43, or the kit according to claim 44 or 45, for use in a method for increasing the efficiency of expression of the artificial nucleic acid molecule in skin tissue, skin cells or a skin cell line, the (pharmaceutical) composition or kit comprising at least one artificial nucleic acid molecule according to any one of claims 7 to 30.

51. The artificial nucleic acid molecule, preferably RNA, according to any one of claims 8 to 30, the (pharmaceutical) composition or vaccine according to any one of claims 31 to 43, or the kit according to claim 44 or 45, for use in a method for increasing the efficiency of expression of the artificial nucleic acid molecule in a muscle tissue, a muscle cell or a muscle cell line, the (pharmaceutical) composition or kit comprising at least one artificial nucleic acid molecule according to any one of claims 8 to 30.

52. A method of treating or preventing a disease optionally selected from the group consisting of a genetic disease, a cancer, an infectious disease, an inflammatory disease, (auto) immune disease, an allergy and/or for gene therapy and/or immunomodulation, wherein the method comprises administering to a subject in need thereof an effective amount of an artificial nucleic acid molecule, preferably RNA, according to any of claims 1 to 30, a (pharmaceutical) composition or vaccine according to any of claims 31 to 43, or a kit according to any of claims 44 or 45.

53. A method for increasing the efficiency of expression of an artificial nucleic acid molecule, preferably RNA, comprising at least one coding region encoding a protein or peptide, preferably a protein or peptide according to any one of claims 11 to 16, comprising

(a) Binding said coding region to at least one 5 'UTR element derived from the 5' UTR of a gene selected from HSD17B4, ASAH1, ATP5a1, MP68, ndifa 4, NOSIP, RPL31, SLC7A3, TUBB4B, and UBQLN2, or a corresponding RNA sequence, homologue, fragment, or variant thereof;

(b) binding said coding region to at least one 3 'UTR element derived from the 3' UTR of a gene selected from PSMB3, CASP1, COX6B1, GNAS, ndifa 1, and RPS9, or a corresponding RNA sequence, homolog, fragment, or variant thereof; and

(c) obtaining an artificial nucleic acid molecule, preferably RNA, according to any one of claims 1 to 30.

54. A method of identifying a combination of a 5 'UTR and a 3' UTR capable of increasing expression efficiency in a desired tissue or a cell derived from a desired tissue, comprising:

a) generating a library of artificial nucleic acid molecules ("test constructs"), each comprising a "reporter ORF" encoding a detectable reporter polynucleotide, preferably a selected luciferase or eGFP, operably linked to one of the 5 'UTRs and/or one of the 3' UTRs as defined in claim 3;

b) providing an artificial nucleic acid molecule comprising a "reporter ORF" operably linked to a reference 5 'UTR and a 3' UTR, preferably RPL32 and ALB7, as a "reference construct";

c) introducing the test construct and the reference construct into a desired tissue or cell under suitable conditions to allow expression thereof;

d) detecting and quantifying expression of the polypeptides from the "reporter ORFs" of the test construct and the reference construct;

e) comparing polypeptide expression of the test construct and the reference construct;

wherein a test construct characterized by increased expression of the polypeptide as compared to a reference construct is identified as capable of increasing expression efficiency in a desired tissue or cell.

Drawings

FIG. 1: average expression profile of selected (poly) peptides and proteins of interest of RNA constructs comprising the UTR combinations of the present invention.

FIG. 2: average expression profile of RNA constructs comprising a UTR combination of the invention operably linked to coding regions encoding different (poly) peptides or proteins of interest and a64 poly a sequence followed by N5 as 3' UTR.

FIG. 3: the average expression profile of RNA constructs comprising polycytidylic acid and histone stem loops in different cell lines in addition to the UTR combinations of the present invention operably linked to coding regions encoding different (poly) peptides or proteins of interest.

FIG. 4: average expression profiles in different cell lines of RNA constructs comprising a combination of UTRs of the invention operably linked to a coding region encoding Erythropoietin (EPO).

FIG. 5: average expression profile in Human Diploid Fibroblasts (HDFs) of RNA constructs comprising a combination of UTRs of the invention operably linked to coding regions encoding different (poly) peptides or proteins of interest.

FIG. 6: average expression profiles in different cell lines of RNA constructs comprising a combination of UTRs of the invention operably linked to a coding region of an antigenic construct encoding a protein of interest.

FIG. 7: average expression profile in HeLa cells of RNA constructs comprising a combination of UTRs of the invention operably linked to coding regions encoding different (poly) peptides or proteins of interest.

FIG. 8: average expression profile in HepG2 cells of RNA constructs comprising a combination of UTRs of the invention operably linked to coding regions encoding different (poly) peptides or proteins of interest.

FIG. 9: average expression profile in HSkMC cells of RNA constructs comprising a combination of UTRs of the invention operably linked to coding regions encoding different (poly) peptides or proteins of interest.

FIG. 10: average expression profiles in different cell lines of RNA constructs comprising a combination of UTRs of the invention operably linked to a coding region encoding rabies virus glycoprotein (RAVG).

FIG. 11: average expression profile in HEK293T cells of RNA constructs comprising a combination of UTRs of the invention operably linked to coding regions encoding different (poly) peptides or proteins of interest.

Examples

In the following, specific examples are presented illustrating various embodiments and aspects of the invention. However, the scope of the invention should not be limited by the particular embodiments described herein. The following preparations and examples are given to enable those skilled in the art to more clearly understand and practice the present invention. The scope of the invention is not limited, however, to the exemplary embodiments, which are intended as illustrations of only a single aspect of the invention, and functionally equivalent methods are within the scope of the invention. Indeed, various modifications of the invention in addition to those described herein will become apparent to those skilled in the art from the foregoing description, the accompanying drawings, and the following examples. All such modifications are intended to fall within the scope of the appended claims.