Synthèse et propriétés d’ARNs modifiés en position 2’ via des ponts disulfures

Abstract : RNAs are involved in numerous biological processes and can adopt different secondary structures. Thanks to their properties, they are powerful biological tools for diverse applications, such as small interfering RNA (siRNA) for gene silencing. Modified RNAs have proven to be essential to improve their properties, and to facilitate the study of their biological and therapeutic functions.This manuscript reports the synthesis and properties of 2’-O-modified RNAs bearing disulfide-containing groups, sensitive to reductive environment.The first part describes the synthesis of siRNAs prodrugs bearing lipophilic benzyldithiomethyl groups. The thermal stability, the serum stability and the response to glutathione treatment of modified siRNAs are thoroughly investigated. The gene silencing and the gymnotic delivery of several siRNAs are assessed, and demonstrates promising results on Ewing’s sarcoma cell line.A second part concerns the co-delivery of siRNAs and a hydrophobic anti-cancer drug (doxorubicin) using a self-immolative spacer bearing disulfide bonds. The chemico-physical properties of these conjugates are determined and the recovery of native siRNA and doxorubicin in response to reductive treatment is highlighted.A third part presents the conjugation of RNAs to small molecules (sugars, coumarin, biotin, deoxycholic acid, glutathione) using disulfide linkages. The synthesis of the RNA conjugates and their release in reducing conditions are also demonstrated.The last part reports the synthesis and the impact of an intrastrand dimethylene disulfide bridge between 2’-O-positions of two adjacent nucleotides in an RNA duplex and in the loop of RNA hairpins. Then, the influence of this linkage on the folding of a biologically relevant RNA structure is reported. Finally, an application of a constrained hairpin as a fluorescent molecular beacon highlights its potential use in tools for understanding RNA folding and in probes for the detection of reducing reagents
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Florian Gauthier. Synthèse et propriétés d’ARNs modifiés en position 2’ via des ponts disulfures. Biologie moléculaire. Université Montpellier, 2018. Français. ⟨NNT : 2018MONTS120⟩. ⟨tel-02356550⟩

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