基于糖代谢重编程重塑肿瘤微环境探讨紫草素调控小细胞肺癌免疫应答的机制研究

Pd-1/pd-l checkpoint inhibitors have made a breakthrough in the treatment of small-cell lung cancer(SCLC), but the respond is still low and the benefit time is short, which may be closely related to macrophages infiltrating in the tumor microenvironment.M1 macrophages obtain energy through anaerobic glycolysis, while M2 macrophages obtain energy through aerobic glycolysis. Changes in glucose metabolism patterns affect different polarization states and functions of macrophages. Studies have found that macrophages in tumor microenvironment can be remodeled, and the transformation from M2 to M1 can be induced by specific means or drugs to reshape tumor immune microenvironment. Our preliminary results showed shikonin change glucose metabolism reprogramming in SCLC cells, combined with the literature and previous work, we propose a hypothesis: shikonin can change glucose metabolism reprogramming in SCLC, prompting the macrophage polarization, converting M2 to M1, regulating immune microenvironment, increasing immune response. C-myc is the key gene. In this study, the macrophages phenotypic and indicators of glucose metabolism would be detected before and after shikonin intervention at the cellular and animal levels. The results will be helpful to clarify the mechanism of shikonin in remodeling tumor immune microenvironment based on glycometabolism reprogramming, and provide new research ideas for TCM to improve immune response.

免疫治疗在小细胞肺癌中取得了突破性进展，但有效率仍较低，获益时间短，研究表明这可能与浸润在肿瘤微环境中的巨噬细胞密切相关。M1 型巨噬细胞是通过无氧糖酵解途径获取能量，M2 型巨噬细胞则通过有氧糖酵解途径获取能量，糖代谢方式的改变影响着巨噬细胞的不同极化状态及其功能。研究发现通过特定的手段或药物，诱导M2型向M1 型转化，可重塑肿瘤免疫微环境。我们前期发现紫草素有改变癌细胞糖代谢重编程的作用，结合文献和前期工作，我们提出假说：紫草素可改变小细胞肺癌细胞糖代谢重编程，进而促使巨噬细胞发生极化，M2型转化为M1型，重塑小细胞肺癌免疫微环境，小细胞肺癌提高免疫应答。C-Myc是代谢调控免疫的关键基因。本项目通过在细胞和动物水平紫草素可改变癌细胞代谢重编程，检测巨噬细胞的表型转化。研究结果有助于明确紫草素从糖代谢重编程角度重塑肿瘤免疫微环境的作用及机制，为中医药提高免疫治疗应答提供新的研究思路。

PD-1/PD-L1免疫检查点抑制剂为代表的免疫治疗的应答率和应答时间很大程度依赖于肿瘤细胞和肿瘤微环境免疫调节的相互作用，通过调控肿瘤细胞的代谢重编程有望重塑肿瘤微环境，提高免疫治疗疗效。紫草素（shikonin）是从中药紫草中提取的萘醌类化和物，具有抗肿瘤、抗炎、抗氧化、免疫调节等多种药理活性。我们前期发现紫草素有改变小细胞肺癌细胞糖代谢重编程的作用。本项目通过体内和体外实验，以人小细胞肺癌细胞株为模型，研究紫草素对小细胞肺癌细胞恶性生物学行为和代谢相关通路的影响，利用二代高通量测序分析基因表达差异，对差异基因进行GO和KEGG富集分析，并利用real-time PCR验证挑选的差异基因。研究结果显示ATF3基因与小细胞肺癌细胞生存和凋亡密切有关，是参与调控小细胞肺癌细胞恶性表型的关键基因。ATF3是铁死亡调控的关键转录因子，研究结果表明ATF3通过抑制细胞中活性氧ROS和还原型谷胱甘肽GSH含量等代谢，诱导铁死亡的发生。紫草素通过促进ATF3的表达，诱导铁死亡的发生，进而影响小细胞肺癌细胞的恶性进展。体内实验验证紫草素通过诱导铁死亡调控细胞恶性表型延缓小细胞肺癌的恶性进展。本课题有助于明确紫草素从调控肿瘤代谢角度重塑肿瘤免疫微环境的作用及机制，为拓宽紫草素的临床应用范围奠定实验基础，并为中医药提高免疫治疗应答提供新的研究思路。