Click-modified mrna
AssigneeBaseclick GmbH
InventorsThomas FRISCHMUTH, Sascha SERDJUKOW, Birgit GRAF, Stefano CROCE
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Abstract
The present invention relates to alkyne- and/or azide-modified mRNA, processes for producing such modified mRNA, cells which are transfected to include the modified mRNA, pharmaceutical compositions containing the modified mRNA or cells including the modified mRNA, and to uses of such mRNA, cells or pharmaceutical compositions in mRNA based therapeutic and/or prophylactic applications.
Claims25
Modified messenger RNA (mRNA), comprising a 5′-cap structure, a 5′-untranslated region (5′-UTR), an open reading frame region (ORF), a 3′-untranslated region (3′-UTR) and a poly(A) tail region, characterized in that it contains at least one of an alkyne- or azide-modification in at least one nucleotide within at least one of the ORF, the 5′-UTR, the 3′-UTR and the poly(A) tail region.
Modified mRNA according to any one of the preceding claims, wherein at least one of the four standard types of nucleotides is present in modified form compared to the non-modified form in a ratio of 1:100 to 10:1, preferably 1:10 to 1:10 or 1:1.
Modified mRNA according to any one of the preceding claims, characterized in that it contains otherwise modified natural or artificial nucleotides, preferably pseudouridine or N1-methylpseudouridine.
Modified mRNA according to any one of the preceding claims, wherein the modified mRNA contains one or more of a detectable label and a functional molecule introduced via a click reaction of the modified mRNA with a correspondingly modified alkyne- or azide-containing detectable label or functional molecule.
Process for the production of the modified mRNA according to any one of the preceding claims, wherein the process comprises in vitro transcribing mRNA from a DNA template or alternatively performing a fermentation process using prokaryotic or eukaryotic host cells to express a DNA template contained in an expression vector wherein the process is performed in the presence of an RNA polymerase and a nucleotide mixture containing the four standard types of nucleotides required for mRNA transcription, in which nucleotide mixture at least a part of at least one of the four types of nucleotides is modified to contain an alkyne- or azide-modification.
Process for the production of a modified mRNA containing an alkyne- or an azide-modification at the poly(A) tail, wherein the process comprises performing a poly(A) polymerase addition reaction at the poly(A) tail on an mRNA in the presence of ATP, wherein ATP is at least partly alkyne- or azide-modified at the adenosine.
Method for stabilizing RNA, especially mRNA, wherein an alkyne- and/or an azide-modification is introduced by including at least one of the four standard types of nucleotides (ATP, CTP, GTP and UTP) in partly or completely alkyne- and/or azide-modified form during RNA synthesis and/or in a poly(A) polymerase addition reaction to produce a modified (m)RNA, and optionally one or more of a detectable label and a functional molecule are introduced via a click reaction of the modified (m)RNA with a correspondingly modified alkyne- or azide-containing detectable label or functional molecule.
Dependencies are inferred from the claim text.