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Structure–Activity Relationship Studies on the Immune Stimulatory Effects of Base‐Modified CpG Toll‐Like Receptor 9 Agonists
Author(s) -
Jurk Marion,
Kritzler Andrea,
Debelak Harald,
Vollmer Jörg,
Krieg Arthur M.,
Uhlmann Eugen
Publication year - 2006
Publication title -
chemmedchem
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.817
H-Index - 100
eISSN - 1860-7187
pISSN - 1860-7179
DOI - 10.1002/cmdc.200600064
Subject(s) - tlr9 , toll like receptor 9 , guanine , hypoxanthine , oligonucleotide , cpg site , cytosine , chemistry , receptor , stimulation , biochemistry , purine , cpg oligodeoxynucleotide , microbiology and biotechnology , dna , biology , stereochemistry , dna methylation , gene , nucleotide , gene expression , enzyme , neuroscience
Synthetic oligodeoxynucleotides containing unmethylated deoxycytidylyl‐deoxyguanosine dinucleotide (CpG) motifs are able to stimulate potent immune responses through a signaling pathway involving Toll‐like receptor 9 (TLR9). We have investigated the structure–activity relationship (SAR) of base‐modified CpG oligonucleotides with TLR9 by measuring TLR9 activation by 20‐mer oligonucleotides having just a single human recognition motif (5′‐GTCGTT‐3′) in functional cell‐based TLR9 assays. Substitution of guanine by hypoxanthine and 6‐thioguanine resulted in activity similar to the unmodified parent molecule, whereas purine, 2‐aminopurine, 2,6‐diaminopurine, and 8‐oxo‐7,8‐dihydroguanine substitution resulted in approximately 40–60 % reduction in activity, and 7‐deazaguanine substitution led to the strongest (80 %) reduction in TLR9 stimulation. Furthermore, none of the investigated modifications at C5 and N4 of cytosine were well tolerated with respect to human TLR9 stimulation. Our results are compatible with a SAR model in which guanine is recognized by the Hoogsteen site, and C5 is most critical for recognition of cytosine. In addition, we found significant species‐specific differences between human and murine TLR9 recognition, which demonstrates the importance of choosing appropriate assay systems for SAR studies.