Patent ID: 8084015

Claim:
A process for preparing a stabilized immunostimulatory complex comprising a cationic peptide immunogen and anionic CpG oligonucleotide comprising the steps of: (a) obtaining a peptide immunogen that has a positive charge at a pH in the range of 5.0 to 8.0, wherein the charge is determined by assigning a +1 charge for each lysine (K), arginine (R) or histidine (H), a −1 charge for each aspartic acid (D) or glutamic acid (E) and a charge of 0 for all other amino acids in the peptide immunogen, and wherein the charge of the peptide immunogen can be adjusted by adding to its N-terminal or C-terminal, amino acid selected from the group consisting of lysine, arginine, histidine, and a mixture thereof; (b) obtaining a CpG oligonucleotide that has a negative charge at a pH in the range of 5.0 to 8.0, wherein the CpG oligonucleotide is a single-stranded DNA comprising 8 to 64 nucleotide bases with a repeat of a cytosine-guanidine motif and the number of repeats of the CpG motif is in the range of 1 to 10, and wherein the charge is determined by assigning a −1 charge for each phosphodiester or phosphorothiorate group and the CpG oligonucleotide can be modified with a phosphorothiorate group at the 5′ end; (c) dissolving or dispersing the cationic peptide immunogen in an aqueous phase selected from the group consisting of distilled deionized water, saline, PBS and a mixture thereof with the proviso that the pH of the aqueous phase is lower than the ionization point of the peptide immunogen; (d) dissolving the anionic CpG oligonucleotide in an aqueous phase selected from the group consisting of distilled deionized water, saline, PBS and a mixture thereof; (e) adding the CpG oligonucleotide in the aqueous phase dropwise to the solution or dispersion of the cationic peptide immunogen in an amount to form a stabilized immunostimulatory complex in the form of particles of the peptide immunogen and the CpG oligonucleotide in a charge ratio of the cationic peptide immunogen to the CpG oligonucleotide in the range of 16:1 to 1:1.