Lnp compositions comprising rna and methods for preparing, storing and using the same

A method of preparing a composition comprising LNPs dispersed in a final aqueous phase, wherein the LNPs comprise a cationically ionizable lipid and RNA;

• the final aqueous phase comprises a final buffer system comprising a final buffer substance and a final monovalent anion, the final buffer substance being selected from the group consisting of Tris and its protonated form, Bis-Tris-methane and its protonated form, and TEA and its protonated form, and the final monovalent anion being selected from the group consisting of chloride, acetate, glycolate, lactate, the anion of MES, the anion of MOPS, and the anion of HEPES;
• the concentration of the final buffer substance in the composition is at most about 25 mM;
• and the final aqueous phase is substantially free of inorganic phosphate anions, substantially free of citrate anions, and substantially free of anions of EDTA;
• wherein the method comprises:
• (I) preparing a formulation comprising LNPs dispersed in the final aqueous phase, wherein the LNPs comprise the cationically ionizable lipid and RNA;
• and
• (II) optionally freezing the formulation to about −10° C. or below,
• thereby obtaining the composition,
• wherein step (1) comprises:
• (a) preparing an RNA solution containing water and a first buffer system;
• (b) preparing an ethanolic solution comprising the cationically ionizable lipid and, if present, one or more additional lipids;
• (c) mixing the RNA solution prepared under (a) with the ethanolic solution prepared under (b), thereby preparing a first intermediate formulation comprising the LNPs dispersed in a first aqueous phase comprising the first buffer system;
• and
• (d) filtrating the first intermediate formulation prepared under (c) using a final aqueous buffer solution comprising the final buffer system,
• thereby preparing the formulation comprising the LNPs dispersed in the final aqueous phase.