title: Bioconjugation of Small Molecules to RNA Impedes Its Recognition by Toll-Like Receptor 7.
abstract: A fundamental mechanism of the innate immune system is the recognition,viaextra- and intracellular pattern-recognition receptors, of pathogen-associated molecular patterns. A prominent example is represented by foreign nucleic acids, triggering the activation of several signaling pathways. Among these, the endosomal toll-like receptor 7 (TLR7) is known to be activated by single-stranded RNA (ssRNA), which can be specifically influenced through elements of sequence structure and posttranscriptional modifications. Furthermore, small molecules TLR7 agonists (smTLRa) are applied as boosting adjuvants in vaccination processes. In this context, covalent conjugations between adjuvant and vaccines have been reported to exhibit synergistic effects. Here, we describe a concept to chemically combine three therapeutic functions in one RNA bioconjugate. This consists in the simultaneous TLR7 stimulation by ssRNA and smTLRa as well as the therapeutic function of the RNA itself, e.g., as a vaccinating or knockdown agent. We have hence synthesized bioconjugates of mRNA and siRNA containing covalently attached smTLRa and tested their function in TLR7 stimulation. Strikingly, the bioconjugates displayed decreased rather than synergistically increased stimulation. The decrease was distinct from the antagonistic action of an siRNA bearing a Gm motive, as observed by direct comparison of the effects in the presence of otherwise stimulatory RNA. In summary, these investigations showed that TRL7 activation can be impeded by bioconjugation of small molecules to RNA.
authors.affiliation: Institute of Pharmacy and Biochemistry, Johannes Gutenberg-University Mainz , Mainz , Germany.
authors.forename: Isabell
authors.initials: I
authors.lastname: Hellmuth
authors.affiliation: Department of Infectious Diseases, Medical Microbiology and Hygiene, University of Heidelberg , Heidelberg , Germany.
authors.forename: Isabel
authors.initials: I
authors.lastname: Freund
authors.affiliation: Department of Dermatology, University Medical Center of the Johannes Gutenberg-University Mainz , Mainz , Germany.
authors.forename: Janine
authors.initials: J
authors.lastname: Schlöder
authors.affiliation: Institute of Pharmacy and Biochemistry, Johannes Gutenberg-University Mainz , Mainz , Germany.
authors.forename: Salifu
authors.initials: S
authors.lastname: Seidu-Larry
authors.affiliation: Institute of Pharmacy and Biochemistry, Johannes Gutenberg-University Mainz , Mainz , Germany.
authors.forename: Kathrin
authors.initials: K
authors.lastname: Thüring
authors.affiliation: Institute of Pharmacy and Biochemistry, Johannes Gutenberg-University Mainz , Mainz , Germany.
authors.forename: Kaouthar
authors.initials: K
authors.lastname: Slama
authors.affiliation: Institute of Organic Chemistry, Johannes Gutenberg-University Mainz , Mainz , Germany.
authors.forename: Jens
authors.initials: J
authors.lastname: Langhanki
authors.affiliation: Max Planck Institute for Polymer Research (MPG) , Mainz , Germany.
authors.forename: Stefka
authors.initials: S
authors.lastname: Kaloyanova
authors.affiliation: Department of Infectious Diseases, Medical Microbiology and Hygiene, University of Heidelberg , Heidelberg , Germany.
authors.forename: Tatjana
authors.initials: T
authors.lastname: Eigenbrod
authors.affiliation: Institute of Organic Chemistry, Johannes Gutenberg-University Mainz , Mainz , Germany.
authors.forename: Matthias
authors.initials: M
authors.lastname: Krumb
authors.affiliation: Max Planck Institute for Polymer Research (MPG) , Mainz , Germany.
authors.forename: Sandra
authors.initials: S
authors.lastname: Röhm
authors.affiliation: Max Planck Institute for Polymer Research (MPG) , Mainz , Germany.
authors.forename: Kalina
authors.initials: K
authors.lastname: Peneva
authors.affiliation: Institute of Organic Chemistry, Johannes Gutenberg-University Mainz , Mainz , Germany.
authors.forename: Till
authors.initials: T
authors.lastname: Opatz
authors.affiliation: Department of Dermatology, University Medical Center of the Johannes Gutenberg-University Mainz , Mainz , Germany.
authors.forename: Helmut
authors.initials: H
authors.lastname: Jonuleit
authors.affiliation: Department of Infectious Diseases, Medical Microbiology and Hygiene, University of Heidelberg , Heidelberg , Germany.
authors.forename: Alexander H
authors.initials: AH
authors.lastname: Dalpke
authors.affiliation: Institute of Pharmacy and Biochemistry, Johannes Gutenberg-University Mainz , Mainz , Germany.
authors.forename: Mark
authors.initials: M
authors.lastname: Helm
keywords: bioconjugate; click chemistry; immunostimulation; mRNA; siRNA; small molecules; toll-like receptor