Structure-efficacy relationships of immunostimulatory activity of CpG-containing oligodeoxynucleotides on mouse spleen cells.
OBJECTIVE
To study the relationship between primary structures of oligodeoxynucleotides (ODN) containing unmethylated deoxycytidyldeoxyguanosine (CpG) dinucleotide motifs and their immunostimulatory activities in mouse spleen cells.


METHODS
A series of CpG ODN with different primary structures were synthesized. Their capabilities to stimulate mouse spleen cell proliferation were determined by [3H]thymidine incorporation assay. Cytokine (interleukin [IL]-6, IL-12, and IFN-alpha) secretion spectra induced by CpG ODN were assessed by ELISA. The ability of CpG ODN to activate natural killer cells was evaluated by standard 4 h (51)Cr-release assay. Flow cytometry was utilized to examine the expressions of various lymphocyte surface molecules on diverse immunocytes. An effective CpG ODN for murine, ODN1826, was set as the template of modification and the positive control.


RESULTS
The immunostimulatory activities of CpG ODN with different sequences and compositions varied markedly, both in character and in extent. It was useless for improving the immunostimulatory activity of ODN1826 by simply increasing the functional hexameric CpG motif number, modifying the site of CpG motifs, or changing the distance between multi-CpG motifs. However, an addition of a self-complementary palindrome structure at the 3'-end, but not the 5'-end of CpG ODN, aroused marked improvement in its activity. Several designed ODN had superior comprehensive immunostimulatory properties compared to ODN1826.


CONCLUSIONS
The immunostimulatory activity of a CpG ODN was relevant to its primary structure. It was useless for promoting immunostimulatory activity to simply change CpG motif number, space, or distance. The 3'-end palindrome structure of CpG ODN is associated with enhanced immunostimulatory activity.
Du
Hai-Yan
HY
Department of Pharmacology and Toxicology, Beijing Institute of Radiation Medicine, Beijing 100850, China. songhf@nic.bmi.ac.cn.
Xia
Shao-You
SY
Song
Hai-Feng
HF
Li
Na
N
Shang
Ming-Mei
MM
Zou
Jia
J
Wang
Qing-Qing
QQ
Ou
Lun
L
Sun
Xiao
X
Ji
Ai-Guo
AG
Tang
Zhong-Ming
ZM
D000276:Adjuvants, Immunologic / Q000737:chemistry / Q000494:pharmacology*; D000818:Animals; D001402:B-Lymphocytes / Q000187:drug effects / Q000276:immunology / Q000378:metabolism; D018122:B7-1 Antigen / Q000378:metabolism; D001483:Base Sequence; D049109:Cell Proliferation / Q000187:drug effects*; D002478:Cells, Cultured; D018899:CpG Islands / Q000235:genetics; D016207:Cytokines / Q000378:metabolism; D003602:Cytotoxicity, Immunologic / Q000187:drug effects; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D007694:Killer Cells, Natural / Q000187:drug effects / Q000276:immunology / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D051920:NK Cell Lectin-Like Receptor Subfamily D / Q000378:metabolism; D009838:Oligodeoxyribonucleotides / Q000737:chemistry / Q000235:genetics / Q000494:pharmacology*; D013154:Spleen / Q000166:cytology*; D013329:Structure-Activity Relationship
D000276:Adjuvants, Immunologic; D018122:B7-1 Antigen; C408982:CPG-oligonucleotide; D016207:Cytokines; D051920:NK Cell Lectin-Like Receptor Subfamily D; D009838:Oligodeoxyribonucleotides