Augmented Induction of CD8+ Cytotoxic T cell Response and Antitumor Resistance by Thl-inducing peptide

效应CD8^+ T细胞识别肿瘤细胞中表达的MHC I类结合改变的自身肽。虽然CD4^+ Th1细胞在调节CD8^+ T细胞中的作用已被证实，但其靶表位和抗肿瘤反应中的功能影响仍不清楚。我们研究了结核分枝杆菌（M.）诱导Thl免疫的强效免疫原性肽是否有助于CD8^+ T细胞的产生和对不相关的肿瘤特异性抗原的保护性抗肿瘤免疫反应。结核分枝杆菌Ag85B的主要Th表位Peptide-25优先诱导C57BL/6小鼠的CD4^+ Thl细胞，并对共免疫的不相关抗原肽的Thl生成有增强作用。用peptide -25和OVA或peptide -25和B16黑色素瘤肽（TRP-2）共同免疫MHC I类小鼠，分别导致OVA和TRP-2肽特异性CD8^+ T细胞显著增加。这种增强的反应依赖于肽-25特异性CD4^+ I…更多的产生fn -y的Th1细胞。在肿瘤保护实验中，用peptide -25和OVA免疫可增强针对OVA特异性的CD8^+细胞毒性细胞的生成，并抑制表达OVA肽的EL-4胸腺瘤的生长，导致肿瘤排斥反应，这是单独用OVA免疫所不能达到的。肽-25反应性Thl细胞在肽-25存在下抑制树突状细胞（DCs），导致它们激活并向CD8^+细胞毒性T细胞呈递OVA肽。这一结果表明，Peptide-25不仅自身诱导thl应答，而且通过与CD4^+ T细胞的相互作用诱导DCs的thl诱导活化。因此，我们培育了TCR转基因小鼠（P25 TCR- tg），表达肽-25反应性克隆T细胞的TCR a链和β链，并分析了P25 TCR- tg诱导CD4^+ T细胞的Thl发育，以阐明肽-25诱导Thl分化的细胞和分子机制。在中性条件下，在I-A^b脾抗原呈递细胞存在的情况下，来自P25 TCR-Tg的初始CD4^+ T细胞在肽-25刺激下优先发育Thl细胞。即使在抗ifn -y和抗il -12存在的情况下，也观察到肽-25诱导的Thl分化。此外，在缺乏IFN-y或IL-12的情况下，携带I-A^b的中国仓鼠卵巢细胞（Peptide-25-I-A^b-CHO）诱导幼稚的CD4^+ T细胞从P25 TCR-Tg向Thl分化。用Peptide-25-I-A^b-CHO刺激TCR 3 h后，定量RT-PCR观察到瞬时T-bet上调和GATA-3表达抑制，两者均不依赖于IFN-y和IL-12。这些结果表明，肽-25/I-A^b和TCR之间的相互作用可能主要影响初始CD4^+ T细胞向Th1亚群分化的命运。综上所述，这些结果表明肽-25具有强大的佐剂活性，为CTL诱导肿瘤抗原提供了有效的帮助。少

The effector CD8^+ T cells recognize MHC class I binding altered self-peptides expressed in tumor cells. Although the requirement for CD4^+ Th1 cells in regulating CD8^+ T cells has been documented, their target epitopes and functional impact in antitumor responses remain unclear. We examined whether a potent immunogenic peptide of Mycobacterium (M.) tuberculosis eliciting Thl immunity contributes to the generation of CD8^+ T cells and to protective antitumor immune responses to unrelated tumor-specific antigens. Peptide-25, a major Th epitope of Ag85B from M. tuberculosis preferentially induced CD4^+ Thl cells in C57BL/6 mice and showed an augmenting effect on Thl generation for coimmunized unrelated antigenic peptides. Coimmunization of mice with Peptide-25 and OVA or Peptide-25 and B16 melanoma peptide (TRP-2) for MHC class I led to a profound increase in CD8^+ T cells specific for OVA and TRP-2 peptides, respectively. This heightened response depended on Peptide-25 specific CD4^+ I … More FN-y-producing Th1 cells. In tumor protection assays, immunization with Peptide-25 and OVA resulted in the enhancement of CD8^+ cytotoxic cell generation specific for OVA and the growth inhibition of EL-4 thymoma expressing OVA peptide leading to the tumor rejection, that were not achieved by immunization with OVA alone. Peptide-25 reactive Thl cells counteractivated dendritic cells (DCs) in the presence of Peptide-25 leading them to activate and present OVA peptide to CD8^+ cytotoxic T cells. This result indicates that Peptide-25 not only induces a Thl-response by itself but also induces Thl-inducing activation of DCs through interactions with CD4^+ T cells. Therefore, we generated TCR transgenic mice (P25 TCR-Tg) expressing TCR a-and β-chains of Peptide-25-reactive cloned T cells and analyzed Thl development of CD4^+ T cells from P25 TCR-Tg to elucidate cellular and molecular mechanisms of the induction of Thl differentiation by Peptide-25. Naive CD4^+ T cells from P25 TCR-Tg preferentially develop Thl cells upon Peptide-25 stimulation in the presence of I-A^b splenic antigen-presenting cells under neutral conditions. Peptide-25-induced Thl differentiation is observed even in the presence of anti-IFN-y and anti-IL-12. Furthermore, Peptide-25-loaded I-A^b-transfected Chinese hamster ovary cells (Peptide-25-I-A^b-CHO) induce Thl differentiation of naive CD4^+ T cells from P25 TCR-Tg in the A^bsence of IFN-y or IL-12. Three hours after the TCR stimulation with Peptide-25-I-A^b-CHO, transient T-bet up-regulation and suppression of GATA-3 expression were observed by quantitative RT-PCR, both of which were independent of IFN-y and IL-12. These results imply that interaction between Peptide-25/I-A^b and TCR may primarily influence determination of the fate of naive CD4^+ T cells in their differentiation towards the Th1 subset. Taken together, these results indicate that Peptide-25 exerts potent adjuvant activity and provides efficient help for CTL induction against tumor antigen. Less