Methods of making novel pharmaceutical compositions

A method for preparing an aqueous composition comprising one or more particles wherein:

• (a) each particle comprises a complex of at least one double-stranded polyribonucleotide, or a salt or solvate thereof, and at least one water-soluble, linear polyethyleneimine, or a salt and/or solvate thereof, wherein
• (i) the double-stranded polyribonucleotide is polyinosinic-polycytidylic acid [poly(I:C)], or a salt or solvate thereof, wherein
• at least 40% of the poly(I:C) have at least 850 base pairs, and
• at least 50% of poly(I:C) have between 400 and 5000 base pairs;
• and
• between 5% and 60% of poly(I:C) have less than 400 base pairs;
• between 15% and 30% of poly(I:C) have between 400 and 850 base pairs,
• between 10% and 70% of poly(I:C) have between 850 and 5000 base pairs;
• between 0% and 10% of poly(I:C) have more than 5000 base pairs, and
• (ii) the average molecular weight of the water-soluble, linear polyethyleneimine is between 17 and 23 kDa and has a polydispersity index of <1.5;
• (b) at least 90% of the particles have a mono-modal diameter distribution of below 300 nm;
• (c) the one or more particles each have a z-average diameter of 80+/−20 nm measured according to ISO standard 22412:2017, and each particle diameter has a polydispersity index between 0.1 and 0.6;
• (d) the poly(I:C) concentration is at least 0.5 mg/mL;
• (e) the composition has a pH of between 3.0+/−0.2 and an osmolality of between 300 and 310 mOsm/kg;
• and
• (f) the composition has a zeta potential between 35 mV and 50 mV measured according to ISO standard 13099-2:2012,
• wherein the method comprises:
• (i) preparing a first aqueous solution of poly(I:C) or a salt or solvate thereof, and a second aqueous solution of linear polyethyleneimine or a salt or solvate thereof;
• (ii) independently filtering each respective first and second aqueous solution through a filter having a pore diameter of less than or equal to 500 nm thereby forming a corresponding first and second sterilized solution;
• and
• (iii) mixing each resulting first and second sterilized solution in a mixing chamber thereby forming the aqueous composition.