Method for optimized RIG-I ligands

A method for increasing the selectivity for RIG-I of a RIG-I agonist, comprising the steps of:

• (a) providing a double-stranded polyribonucleotide comprising a sense strand with 24 to 30 nucleotides in length and an antisense strand with 24 to 30 nucleotides in length,
• wherein the sense strand, or the antisense strand, or both the sense strand and the antisense strand are selected from SEQ ID NO:
• 10-15, 18-22, 24-35, 44-45, 49-54, 60-62, 159-166, and 206-209,
• wherein the sense strand and the antisense strand form a fully complementary region of at least 24 base pairs with a blunt end at the 5′-end of the sense strand and the 3′-end of the antisense strand;
• and
• wherein the first 24 nucleotides at the 5′-end of the sense strand are ribonucleotides;
• and
• wherein the sense strand has no 2′-o-methyl modification at a ribonucleotide at a position selected from the group consisting of position number 1, 7, 8, 9, and 14, and
• wherein the last 24 nucleotides at 3′-end of the antisense strand are ribonucleotides and
• wherein the antisense strand has in its last 24 nucleotides no 2′-o-methyl modification at a ribonucleotide at a position selected from the group consisting of position 18, 20, and 23;
• wherein all positions are counted from 5′ to 3′;
• (b) identifying whether the polyribonucleotide of step (a) comprises a purine ribonucleotide at a position selected from the group consisting of position number 12, 15, and 20 in the sense strand, and position number 3 and 10 of the antisense strand, and
• (c) introducing at least one 2′-o-methyl modification at a purine ribonucleotide identified in step (b).