基于CaN/NFAT信号通路，疏肝凉血方逆转Foxp3+Treg介导乳腺癌免疫逃逸的药理机制

The Foxp3+ regulatory T cell (Foxp3+Treg) mediated immune suppression is a key mechanism of tumor immune escape. Calcineurin (CaN)/ nuclear factor of activated T cells (NFAT) pathway activation can induce expression of Foxp3 in Treg, which could promote immune escape in breast cancer. Stagnation of Liver-Qi and hot blood is the specific pathogenesis of breast cancer of breast cancer in traditional Chinese medicine, and Shugan liangxue prescription can inhibit angiogenesis of breast cancer via reducing CaN activity, restore T cell immune surveillance function and enhance anti-cancer immunity. However, whether the prescription can regulate Foxp3+Treg function through inhibiting CaN/NFAT pathway is not clear. Therefore, this research proposed the hypothesis that “Shugan-liangxue prescription reverses immune escape of breast cancer via inhibiting CaN/NFAT pathway”. The in vivo chemical-induced rat breast cancer model and in vitro “CD4+CD25+Treg” model were used. The FK506 was used to block CaN/NFAT pathway. In the cellular and molecular levels, the CaN enzyme activity, ratio of NFAT enter nucleus, Foxp3 expression, the T cell subgroup within their cytokines were detected. The correlations between change amplitude of each index and anticancer potency were regressed to confirm the pharmacological effects of Shuganliangxue prescription on different segment of CaN/NFAT pathway, and the underlying mechanisms of this prescription on breast cancer immune escape were explored.

Foxp3+调节性T细胞(Foxp3+Treg)介导的免疫抑制是肿瘤免疫逃逸的关键机制。CaN/NFAT通路过度活化可诱导Treg表达Foxp3,促进乳腺癌免疫逃逸。针对乳腺癌肝郁血热中医特异性病机，疏肝凉血方可抑制CaN活性抗乳腺癌血管新生，并能恢复T细胞免疫监视，增强抗癌免疫。但该方剂是否通过抑制CaN/NFAT通路调节Foxp3+Treg功能尚未明确。因此本课题提出“抑制CaN/NFAT通路，疏肝凉血方减少Treg细胞Foxp3表达，逆转乳腺癌免疫逃逸”的工作假说。采用在体化学诱导性大鼠乳腺癌和离体“CD4+CD25+Treg”细胞模型，以FK506阻断CaN/NFAT通路，检测CaN酶活性、NFAT入核比、Foxp3表达、T细胞亚群及细胞因子水平，回归指标变化幅度同抗癌效能的相关性，分析该方剂抑制CaN/NFAT通路,逆转Foxp3+Treg介导乳腺癌免疫逃逸的药理机制。

乳腺癌是女性常见恶性肿瘤，发病率和死亡率高。Foxp3+调节性T细胞(Treg)介导的免疫抑制效应是肿瘤免疫逃逸的关键机制。CaN/NFAT通路过度活化可诱导Treg表达foxp3，促进乳腺癌免疫逃逸。中医药以疏肝凉血治疗乳腺癌肝郁血热证，能恢复T细胞免疫监视，增强抗癌免疫。本项目以化学物诱导致癌，联合完全弗氏佐剂放大免疫效应，复制大鼠乳腺癌免疫模型；给予疏肝凉血方治疗，发现该方剂抑制癌细胞增殖与肿瘤生长，增加肿瘤间质CD8+T细胞浸润，促进IL-12和IFN-γ分泌，抑制IL-10和IL-4分泌，提高“抗癌/促癌”效能，恢复免疫监视功能；在整体动物与离体细胞模型上，确认了方剂通过抑制CaN酶活性，抑制NFAT通路，减少Treg细胞foxp3表达，逆转乳腺癌免疫逃逸；同时通过转录学分析，发现了方剂通过调节脂质、氨基酸免疫代谢，增强细胞免疫抗癌效应。本项目分析了 “疏肝凉血”逆转乳腺癌免疫逃逸的药理机制，为中医药抗乳腺癌的临床应用奠定了基础。