McCluskie M J, Weeratna R D, Davis H L
Loeb Health Research Institute, Ottawa Hospital, Canada.
Mol Med. 2000 Oct;6(10):867-77.
Synthetic oligodeoxynucleotides (ODN) containing immunostimulatory cytosine-guanine phosphate-linked dinucleotide (CpG) motifs are potent systemic and mucosal adjuvants in mice that have synergistic action with numerous other adjuvants, including alum and cholera toxin (CT). Herein, we evaluate CpG ODN with intranasal (IN) delivery of purified hepatitis B surface antigen (HBsAg), relative to and in combination with CT, Escherichia coli heat labile enterotoxin (LT), the B subunit of CT (CTB), and a nontoxic derivative of LT (LTK63).
BALB/c mice were immunized by IN administration of HBsAg, alone or combined with CT, LT, CTB, or LTK63, and/or CpG ODN, or non-CpG control ODN. In addition, the effect of low-or high-volume administration was assessed, in order to target upper respiratory or entire respiratory tract, respectively. HBsAg-specific systemic (immunoglobulins: IgG, IgG1, IgG2a in plasma) and mucosal (IgA in fecal, lung, vaginal, saliva, and gut samples) humoral responses, as well as cell-mediated immune responses including T-cell proliferation and cytokines (interleukins: IL-4, IL-5; interferon: IFN-gamma) were evaluated.
CpG ODN, CT, and LT augmented anti-HBs titers equally, and more so than did CTB or LTK63. CpG ODN acted synergistically with CT and LT, but not CTB or LTK63 to enhance anti-HBs titers. Nevertheless, CpG ODN induced a more Th1-like response for all combinations, compared with the same formulation without CpG. Strength of induced systemic and mucosal immune responses was better with IN delivery of a large volume. A small volume required multiple administrations and higher doses of antigen and adjuvant for equal results. This suggests that delivery of antigen to the lung and/or diges-tive system is superior to delivery to the nasal cavity.
Our results suggest that the synergy between CpG ODN and native toxins (CT, LT) may depend on their enzymatic activity and that the lack of synergy with nontoxic derivatives (LTB, LTK63) arises, since they do not have enzymatic activity. Because both CT and LT are too toxic for use in humans, it is possible that CpG ODN may be combined with bacterial toxin mutants that retain some enzymatic activity to optimize immune augmentation.
含有免疫刺激性胞嘧啶 - 鸟嘌呤磷酸连接二核苷酸（CpG）基序的合成寡脱氧核苷酸（ODN）是小鼠体内强效的全身和黏膜佐剂，可与包括明矾和霍乱毒素（CT）在内的许多其他佐剂产生协同作用。在此，我们评估经鼻内（IN）递送纯化的乙型肝炎表面抗原（HBsAg）时，CpG ODN相对于CT、大肠杆菌不耐热肠毒素（LT）、CT的B亚基（CTB）以及LT的无毒衍生物（LTK63）单独使用或联合使用的效果。
用单独的HBsAg或与CT、LT、CTB或LTK63和/或CpG ODN或非CpG对照ODN联合经鼻内给药免疫BALB/c小鼠。此外，评估了低剂量或高剂量给药的效果，以分别靶向于上呼吸道或整个呼吸道。评估了HBsAg特异性的全身（血浆中的免疫球蛋白：IgG、IgG1、IgG2a）和黏膜（粪便、肺、阴道、唾液和肠道样本中的IgA）体液反应，以及包括T细胞增殖和细胞因子（白细胞介素：IL - 4、IL - 5；干扰素：IFN - γ）在内的细胞介导免疫反应。
CpG ODN、CT和LT同等程度地提高了抗HBs滴度，且比CTB或LTK63提高得更多。CpG ODN与CT和LT协同作用以提高抗HBs滴度，但与CTB或LTK63无协同作用。然而，与不含CpG的相同制剂相比，CpG ODN在所有组合中均诱导出更类似Th1的反应。经鼻内递送大剂量时诱导的全身和黏膜免疫反应强度更好。小剂量需要多次给药以及更高剂量的抗原和佐剂才能获得相同的结果。这表明将抗原递送至肺和/或消化系统优于递送至鼻腔。
我们的结果表明，CpG ODN与天然毒素（CT、LT）之间的协同作用可能取决于它们的酶活性，而与无毒衍生物（LTB、LTK63）缺乏协同作用是因为它们没有酶活性。由于CT和LT对人类使用毒性太大，因此CpG ODN有可能与保留某些酶活性的细菌毒素突变体联合使用以优化免疫增强效果。
