Fate of Tableted Freeze-Dried siRNA Lipoplexes in Gastrointestinal Environment.

Asad Ur Rehman,Marcela Coelho Silva Ribeiro,Virginie Busignies,Christine Charrueau,Pierre Tchoreloff,Virginie Escriou,Nayara Almeida Lage

doi:10.3390/pharmaceutics13111807

Abstract

The incorporation of siRNA into nanocarriers is mandatory to facilitate its intracellular delivery, as siRNA itself cannot enter cells. However, the incorporation of these nanocarriers into oral, solid dosage forms and their fate in the gastrointestinal environment is yet to be explored. In the present work, the fate of, (i) naked siRNA, (ii) freshly prepared siRNA lipoplexes, and (iii) tableted siRNA lipoplexes, in simulated gastric and intestinal fluids was studied. The siRNA, either released from or protected within the lipoplexes, was quantified by gel electrophoresis and siRNA efficacy was assessed in cell transfection. The freshly prepared lipoplexes kept their siRNA load and transfection efficiency totally preserved during 1 h of incubation in simulated gastric fluid at 37 °C. However, in simulated intestinal fluid, despite no release of siRNA from lipoplexes after 6 h of incubation, gene silencing efficacy was dramatically decreased even after 1 h of exposure. The lipoplexes obtained from tablets efficiently protected siRNA in simulated gastric fluid, thus preserving the gene silencing efficacy, whereas their incubation in simulated intestinal fluid resulted in a marked siRNA release and decreased gene silencing efficacy. These results provided a detailed explanation for understanding the fate of siRNA in gastrointestinal conditions, when simply loaded in lipoplexes or formulated in the form of tablets.

Highlights

Therapeutic RNA interference (RNAi) relies on the principle of silencing diseaserelated genes in a sequence-specific manner
They were tested as freshly prepared siRNA lipoplexes in colloidal suspension, and tableted, freeze-dried siRNA lipoplexes after resuspension, in comparison with naked siRNA in solution (Figure 1)
Oral delivery of siRNAs has been considered ever since the Nobel Prize was awarded to RNA interference in 2006 [6]

Summary

Introduction

Therapeutic RNA interference (RNAi) relies on the principle of silencing diseaserelated genes in a sequence-specific manner. After two decades of research, this promising curative modality has become a reality, since three siRNA therapeutics have been approved, i.e., ONPATTRO® (patisiran, Sanofi Genzyme) in 2018 [1], GIVLAARI® (givosiran, Alnylam Pharmaceuticals) in 2019 [2], both being commercialized, and OXLUMO®. In ONPATTRO® , siRNA is formulated in lipid nanoparticles for the intravenous treatment of hereditary amyloidogenic transthyrethin amyloidosis with polyneuropathy in adults [2]. 1 gene to treat primary hyperoxaluria type 1 [3], are both covalently linked to a ligand enabling their specific delivery to hepatocytes (Gal-NAc-siRNA conjugates), and are administered by subcutaneous injection. The administration of nonconjugated siRNAs necessitates their formulation into vectors in order to achieve cellular delivery, since these negatively charged and water-soluble molecules, Pharmaceutics 2021, 13, 1807. Oral delivery of drugs represents a real challenge since physiological and cellular barriers have to be overcome

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